A significant number of homes in the United States are not connected to a central sewage disposal and treatment system and rely on local septic systems to treat wastewater discharged from the home. Such septic systems typically have a septic tank for receiving the wastewater from the home. The wastewater and the solids entrained in it are deposited in the tank where they are decomposed by the action of bacteria and other micro-organisms. The wastewater itself is eventually discharged into a septic field that surrounds the tank and percolates down through the soil. Grease and similar substances in the wastewater causes the soil of the septic field eventually to coagulate forming in effect a seal, and the wastewater is no longer able to drain away sufficiently quickly for the system to operate properly; an overflow or other dire consequence results. The speed with which such a system failure occurs is dependent on the characteristics and condition of the soil, the amount of wastewater discharged, and the materials found in the wastewater.
It is known in the prior art that fermentation supernatants obtained from the fermentation of certain yeasts, when present in the appropriate concentrations, have the ability to accelerate the natural digestion of biologically available organic compounds present in sewage, sludge, grease, and the like. Examples of such materials are described in U.S. Pat. Nos. 3,635,797, 5,464,766, 5,820,758, 5,879,928, 5,885,950, 5,905,037, and 6,699,391. Typically, the fermentation supernatant with its protein component is combined with other ingredients such as surfactants, buffers, citric and lactic acids, urea, preservatives and the like and then diluted with water to form a protein rich cleaning solution. An example of such a protein rich cleaning solution is ACCELL® cleaning solution, a product containing fermentation supernatant, surfactants, buffers, etc., and available from Advanced BioCatalytics Corporation, of Irvine, Calif., the assignee of the present application. In the ACCELL® cleaning solution, the protein component itself represents about 1.5% by weight of the solution and water represents about 83% by weight. The surfactants, buffers, etc., make up the remainder.
The fermentation supernatant includes a protein component comprised of a variety of proteins produced by an aerobic yeast fermentation process. The aerobic yeast fermentation process is conducted within a reactor having aeration and agitation mechanisms, such as aeration tubes and/or mechanical agitators. The starting materials (liquid growth medium, yeast, sugars, additives) are added to the fermentation reactor and the fermentation is conducted as a batch process. After fermentation, the fermentation product may be subjected to additional procedures intended to increase the yield of proteins produced from the process. Examples of these additional procedures include heat shock of the fermentation product, physical and/or chemical disruption of the yeast cells to release additional polypeptides, lysing of the yeast cells, or other procedures described herein and/or known to those of skill in the art. The yeast cells are removed by centrifugation or filtration to produce a supernatant containing the protein component. Various processes for obtaining the supernatant are disclosed in co-pending application, U.S. Patent Application Publication No. 2005-0245414 (Ser. No. 10/837,312, filed on Apr. 29, 2004), entitled “Improving Surface-Active Properties of Surfactants”, and assigned to the assignee of this application. That co-pending application is hereby incorporated by reference herein.
As used herein, the term “protein component” refers to a mixture of proteins that includes a number of proteins having a molecular weight of between about 100 and about 450,000 daltons, and most preferably between about 500 and about 50,000 daltons, and which, when combined with one or more surfactants, enhances the surface-active properties of the surfactants.
In a first example, the protein component comprises the supernatant recovered from an aerobic yeast fermentation process. Yeast fermentation processes are generally known to those of skill in the art, and are described, for example, in the chapter entitled “Baker's Yeast Production” in Nagodawithana T.W. and Reed G., Nutritional Requirements of Commercially Important Microorganisms, Esteekay Associates, Milwaukee, Wis., pp 90-112 (1998), which is hereby incorporated by reference. Briefly, the aerobic yeast fermentation process is conducted within a reactor having aeration and agitation mechanisms, such as aeration tubes and/or mechanical agitators. The starting materials (e.g., liquid growth medium, yeast, a sugar or other nutrient source such as molasses, corn syrup, or soy beans, diastatic malt, and other additives) are added to the fermentation reactor and the fermentation is conducted as a batch process.
After fermentation, the fermentation product may be subjected to additional procedures intended to increase the yield of the protein component produced from the process. Several examples of post-fermentation procedures are described in more detail below. Other processes for increasing yield of protein component from the fermentation process may be recognized by those of ordinary skill in the art.
Saccharomyces cerevisiae is a preferred yeast starting material, although several other yeast strains may be useful to produce yeast ferment materials used in the compositions and methods described herein. Additional yeast strains that may be used instead of or in addition to Saccharomyces cerevisiae include Kluyveromyces marxianus, Kluyveromyces lactis, Candida utilis (Torula yeast), Zygosaccharomyces, Pichia, Hansanula, and others known to those skilled in the art.
In the first embodiment, saccharomyces cerevisiae is grown under aerobic conditions familiar to those skilled in the art, using a sugar, preferably molasses or corn syrup, soy beans, or some other alternative material (generally known to one of skill in the art) as the primary nutrient source. Additional nutrients may include, but are not limited to, diastatic malt, diammonium phosphate, magnesium sulfate, ammonium sulfate zinc sulfate, and ammonia. The yeast is preferably propagated under continuous aeration and agitation between 30 degrees to 35 degrees C. and at a pH of 4.0 to 6.0. The process takes between 10 and 25 hours and ends when the fermentation broth contains between 4 and 8% dry yeast solids, (alternative fermentation procedures may yield up to 15-16% yeast solids), which are then subjected to low food-to-mass stress conditions for 2 to 24 hours. Afterward, the yeast fermentation product is centrifuged to remove the cells, cell walls, and cell fragments. It is worth noting that the yeast cells, cell walls, and cell fragments will also contain a number of useful proteins suitable for inclusion in the protein component described herein.
In an alternative embodiment, the yeast fermentation process is allowed to proceed until the desired level of yeast has been produced. Prior to centrifugation, the yeast in the fermentation product is subjected to heat-stress conditions by increasing the heat to between 40 and 60 degrees C., for 2 to 24 hours, followed by cooling to less than 25 degrees C. The yeast fermentation product is then centrifuged to remove the yeast cell debris and the supernatant, which contains the protein component, is recovered.
In a further alternative embodiment, the fermentation process is allowed to proceed until the desired level of yeast has been produced. Prior to centrifugation, the yeast in the fermentation product is subjected to physical disruption of the yeast cell walls through the use of a French Press, ball mill, high-pressure homogenization, or other mechanical or chemical means familiar to those skilled in the art, to aid the release of intracellular, polypeptides and other intracellular materials. It is preferable to conduct the cell disruption process following a heat shock, pH shock, or autolysis stage. The fermentation product is then centrifuged to remove the yeast cell debris and the supernatant is recovered.
In a still further alternative embodiment, the fermentation process is allowed to proceed until the desired level of yeast has been produced. Following the fermentation process, the yeast cells are separated out by centrifugation. The yeast cells are then partially lysed by adding 2.5% to 10% of a surfactant to the separated yeast cell suspension (10%-20% solids). In order to diminish the protease activity in the yeast cells, 1 mM EDTA is added to the mixture. The cell suspension and surfactants are gently agitated at a temperature of about 25° to about 35° C. for approximately one hour to cause partial lysis of the yeast cells. Cell lysis leads to an increased release of intracellular proteins and other intracellular materials. After the partial lysis, the partially lysed cell suspension is blended back into the ferment and cellular solids are again removed by centrifugation. The supernatant, containing the protein component, is then recovered.
In a still further alternative embodiment, fresh live Saccharomyces cerevisiae is added to a jacketed reaction vessel containing methanol-denatured alcohol. The mixture is gently agitated and heated for two hours at 60 degrees C. The hot slurry is filtered and the filtrate is treated with charcoal and stirred for 1 hour at ambient temperature, and filtered. The alcohol is removed under vacuum and the filtrate is further concentrated to yield an aqueous solution containing the protein component.
Additional details concerning the fermentation processes and other aspects of the protein component are described in U.S. Patent Application Publication No. 2004-0180411 (Ser. No. 10/799,529, filed Mar. 11, 2004), entitled “Altering Metabolism in Biological Processes,” which is assigned to the assignee of the present application. Still further details concerning these processes and materials are described in the aforementioned U.S. Pat. No. 6,999,391 which is also assigned to the assignee of the present application. This patent application and this patent are hereby incorporated by reference herein.
Among the uses proposed for such protein rich cleaning solutions is the cleaning of septic systems, tanks, and drainage fields. Typically, the protein rich cleaning solution is introduced directly into the targeted material. For example, such a cleaning solution has been continuously introduced into a septic field serving a mobile home community by introducing the solution into a central pumping installation that discharges wastewater into the septic field. In this application, a concentration of about fifteen parts per million of the solution relative to the total water flow through the system was used. U.S. Pat. No. 5,885,950 proposes introducing the cleaning solution directly into the septic tank, and covering the drain field with a further diluted version of the solution and then watering to wash the composition into the drain field. It has also been proposed to treat an individual home's septic system by the application through the plumbing system of a measured dose of the cleaning solution on a weekly basis. In both the latter instances, the treatment has to be periodically repeated, and thus each of these approaches has the disadvantage of requiring the homeowner to do periodic maintenance and add another chore to his schedule, one that can be easily overlooked.